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Highly specific aptamer trap for extremophilic RNA polymerases
Ivan Petushkov1, Andrey Feklistov2, Andrey Kulbachinskiy1
1National Research Center "Kurchatov Institute", Moscow, 123182, Kurchatov Sq. 2, Russia; Institute of Gene Biology, Russian Academy of Sciences, Moscow, 119334, Russia.
Biochimie
|May 17, 2024
Summary
Researchers developed a synthetic DNA aptamer that binds bacterial RNA polymerase (RNAP) with high affinity. This aptamer inhibits RNAP activity by blocking DNA interaction, offering potential for structural studies and drug screening.
Area of Science:
- Molecular Biology
- Biochemistry
- Structural Biology
Background:
- Bacterial RNA polymerase (RNAP) holoenzyme uses σ factors for promoter recognition during initiation.
- During elongation, core RNAP nonspecifically binds nucleic acids, locking DNA and RNA in its cleft.
Purpose of the Study:
- To present a synthetic DNA aptamer targeting bacterial RNAP.
- To characterize the aptamer's binding affinity and inhibitory mechanism.
- To explore potential applications of the aptamer.
Main Methods:
- DNA aptamer synthesis and characterization.
- Binding assays with bacterial RNAP (core and holoenzyme).
- Inhibition assays for transcription elongation.
Main Results:
- A synthetic DNA aptamer was identified that binds RNAP from Deinococcus-Thermus bacteria.
- The aptamer exhibits subnanomolar affinity and forms stable complexes under high ionic strength.
- The aptamer inhibits RNAP activity by blocking DNA template interaction.
Conclusions:
- The aptamer likely binds a conserved site in the RNAP downstream DNA-binding cleft, inducing an inactive conformation.
- This aptamer is a valuable tool for RNAP structural studies, affinity purification, and transcriptional inhibitor screening.
